CYB5D2 enhances HeLa cells survival of etoposide-induced cytotoxicity

Xie, Yanyun; Bruce, Anthony; He, Lizhi; Wei, Fengxiang; Tao, Ling; Tang, Damu

doi:10.1139/o11-004

Cited by 11 publications

(23 citation statements)

References 33 publications

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“…Consistent with these structural similarities, human CYB5D2 was reported to bind heme and deletion of the cyt-b5 domain (amino acid residues 55–98) ablated this interaction [9]. Both proteins are 40.4% identical and 55.8% similar within their cyt-b5 domain [10]. It has been previously shown that deletion of the TM domain in CYB5D2 abolished its perinuclear localization, compromising its survival from etoposide treatment.…”

Section: Introductionmentioning

confidence: 56%

“…GST-agarose was subsequently used for GST removal. The recombinant GST-free CYB5D2 and CYB5D2(D86G) proteins were confirmed by Western blot using our in-house generated anti-CYB5D2 rabbit polyclonal antibody [10].…”

Section: Methodsmentioning

confidence: 97%

“…Construction of vectors expressing amino-terminal HA-tagged CYB5D2, CYB5D2(ΔTM), and CYB5D2(Δcyt-b5) has been previously described [10]. CYB5D2(D86G), CYB5D2(Y73A), CYB5D2(Y79A), CYB5D2(Y127A) and CYB5D2 were inserted in amino-terminal HA- or FLAG-tagged pcDNA3-based vectors.…”

Section: Methodsmentioning

confidence: 99%

“…It has been previously shown that deletion of the TM domain in CYB5D2 abolished its perinuclear localization, compromising its survival from etoposide treatment. Furthermore, deletion of the cyt-b5 domain (amino acid residues 35–134) of CYB5D2 increased the sensitivity of HeLa cells to etoposide toxicity [10].…”

Section: Introductionmentioning

confidence: 99%

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CYB5D2 Requires Heme-Binding to Regulate HeLa Cell Growth and Confer Survival from Chemotherapeutic Agents

Bruce

Rybak

2014

PLoS ONE

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The cytochrome b5 domain containing 2 (CYB5D2; Neuferricin) protein has been reported to bind heme, however, the critical residues responsible for heme-binding are undefined. Furthermore, the relationship between heme-binding and CYB5D2-mediated intracellular functions remains unknown. Previous studies examining heme-binding in two cytochrome b5 heme-binding domain-containing proteins, damage-associated protein 1 (Dap1; Saccharomyces cerevisiae) and human progesterone receptor membrane component 1 (PGRMC1), have revealed that conserved tyrosine (Y) 73, Y79, aspartic acid (D) 86, and Y127 residues present in human CYB5D2 may be involved in heme-binding. CYB5D2 binds to type b heme, however, only the substitution of glycine (G) at D86 (D86G) within its cytochrome b5 heme-binding (cyt-b5) domain abolished its heme-binding ability. Both CYB5D2 and CYB5D2(D86G) localize to the endoplasmic reticulum. Ectopic CYB5D2 expression inhibited cell proliferation and anchorage-independent colony growth of HeLa cells. Conversely, CYB5D2 knockdown and ectopic CYB5D2(D86G) expression increased cell proliferation and colony growth. As PGRMC1 has been reported to regulate the expression and activities of cytochrome P450 proteins (CYPs), we examined the role of CYB5D2 in regulating the activities of CYPs involved in sterol synthesis (CYP51A1) and drug metabolism (CYP3A4). CYB5D2 co-localizes with cytochrome P450 reductase (CYPOR), while CYB5D2 knockdown reduced lanosterol demethylase (CYP51A1) levels and rendered HeLa cells sensitive to mevalonate. Additionally, knockdown of CYB5D2 reduced CYP3A4 activity. Lastly, CYB5D2 expression conferred HeLa cell survival from chemotherapeutic agents (paclitaxel, cisplatin and doxorubicin), with its ability to promote survival being dependent on its heme-binding ability. Taken together, this study provides evidence that heme-binding is critical for CYB5D2 in regulating HeLa cell growth and survival, with endogenous CYB5D2 being required to modulate CYP activities.

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Section: Introductionmentioning

confidence: 56%

Section: Methodsmentioning

confidence: 97%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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CYB5D2 Requires Heme-Binding to Regulate HeLa Cell Growth and Confer Survival from Chemotherapeutic Agents

Bruce

Rybak

2014

PLoS ONE

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“…Human neuferricin is 264 amino acids in length and includes a cleavable signal sequence at its N terminus followed by a heme-binding domain that may involve aspartic acid (D86) instead of tyrosine in heme binding (Bruce and Rybak, 2014). CYB5D2 can be detected as secreted hemoprotein in some cell lines but also colocalized with POR in the ER (Xie et al, 2011; Bruce and Rybak, 2014). Ectopic CYB5D2 expression inhibited cell proliferation and anchorage-independent colony growth of HeLa cells (Bruce and Rybak, 2014).…”

Section: Molecular Biochemical and Cellular Properties Of Maprsmentioning

confidence: 99%

Membrane Associated Progesterone Receptors: Promiscuous Proteins with Pleiotropic Functions – Focus on Interactions with Cytochromes P450

2017

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Membrane-associated progesterone receptors (MAPR) are a group of four rather small, partially homologous proteins, which share a similar non-covalent heme-binding domain that is related to cytochrome b5, a well-known functional interaction partner of microsomal cytochrome P450 (CYP) monooxygenase systems. Apart from their structural similarities the four proteins progesterone membrane component 1 (PGRMC1, also referred to as IZA, sigma-2 receptor, Dap1), PGRMC2, neudesin (NENF) and neuferricin (CYB5D2) display surprisingly divergent and multifunctional physiological properties related to cholesterol/steroid biosynthesis, drug metabolism and response, iron homeostasis, heme trafficking, energy metabolism, autophagy, apoptosis, cell cycle regulation, cell migration, neural functions, and tumorigenesis and cancer progression. The purpose of this mini-review is to briefly summarize the structural and functional properties of MAPRs with particular focus on their interactions with the CYP system. For PGRMC1, originally identified as a non-canonical progesterone-binding protein that mediates some immediate non-genomic actions of progesterone, available evidence indicates mainly activating interactions with steroidogenic CYPs including CYP11A1, CYP21A2, CYP17, CYP19, CYP51A1, and CYP61A1, while interactions with drug metabolizing CYPs including CYP2C2, CYP2C8, CYP2C9, CYP2E1, and CYP3A4 were either ineffective or slightly inhibitory. For the other MAPRs the evidence is so far less conclusive. We also point out that experimental limitations question some of the previous conclusions. Use of appropriate model systems should help to further clarify the true impact of these proteins on CYP-mediated metabolic pathways.

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CYB5D2 displays tumor suppression activities towards cervical cancer

Xie¹,

Shen²,

Kapoor³

et al. 2016

Biochimica et Biophysica Acta (BBA) - Molecular Basis of Diseas

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Cervical cancer is caused by infections with human papillomaviruses (HPV) and genetic alternations in the cervical epithelium. While the former is well studied, the latter remains unclear. We report here that CYB5D2/Neuferricin possesses tumor suppressing activity towards cervical tumorigenesis. Ectopic expression of CYB5D2 did not affect HeLa cell proliferation and the cell's ability to form xenograft tumors, but significantly inhibited HeLa cell invasion in vitro and the cell-produced lung metastasis in NOD/SCID mice. Knockdown of CYB5D2 enhanced HeLa cell invasion. Two mutations in CYB5D2, the substitutions of arginine (R) 7 with either proline (P) or glycine (G), were reported in colon cancer. Both CYB5D2(R7P) and CYB5D2(R7G) were incapable of inhibiting HeLa cell invasion. CYB5D2 binds heme, in which aspartate (D) 86 is required. While CYB5D2(D86G) is heme-binding defective, it inhibited HeLa cell invasion. On the other hand, CYB5D2(R7P) and CYB5D2(R7G) bound heme but did not inhibit HeLa cell invasion. Collectively, CYB5D2 inhibits HeLa cell invasion independently of its heme binding. Furthermore, immunohistochemistry examination of CYB5D2 expression in 20 normal cervical tissues and 40 cervical squamous cell carcinomas (SCC) revealed a CYB5D2 reduction in 87.5% (35/40) of SCC. Analysis of CYB5D2 gene expression and genomic alteration data available from Oncomeine™ detected significant reductions of CYB5D2 mRNA in 40 SCCs and CYB5D2 gene copy number in 107 SCCs. Collectively, we provide evidence that CYB5D2 is a candidate tumor suppressor of cervical tumorigenesis.

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CYB5D2 enhances HeLa cells survival of etoposide-induced cytotoxicity

Cited by 11 publications

References 33 publications

CYB5D2 Requires Heme-Binding to Regulate HeLa Cell Growth and Confer Survival from Chemotherapeutic Agents

CYB5D2 Requires Heme-Binding to Regulate HeLa Cell Growth and Confer Survival from Chemotherapeutic Agents

Membrane Associated Progesterone Receptors: Promiscuous Proteins with Pleiotropic Functions – Focus on Interactions with Cytochromes P450

CYB5D2 displays tumor suppression activities towards cervical cancer

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